title: Regulation of the DNA damage response and gene expression by the Dot1L histone methyltransferase and the 53Bp1 tumour suppressor.
abstract: BACKGROUND
  Dot1L, a histone methyltransferase that targets histone H3 lysine 79 (H3K79), has been implicated in gene regulation and the DNA damage response although its functions in these processes remain poorly defined.
  
  
  RESULTS
  Using the chicken DT40 model system, we generated cells in which the Dot1L gene is disrupted to examine the function and focal recruitment of the 53Bp1 DNA damage response protein. Detailed kinetic and dose response assays demonstrate that, despite the absence of H3K79 methylation demonstrated by mass spectrometry, 53Bp1 focal recruitment is not compromised in these cells. We also describe, for the first time, the phenotypes of a cell line lacking both Dot1L and 53Bp1. Dot1L⁻/⁻ and wild type cells are equally resistant to ionising radiation, whereas 53Bp1⁻/⁻/Dot1L⁻/⁻ cells display a striking DNA damage resistance phenotype. Dot1L and 53Bp1 also affect the expression of many genes. Loss of Dot1L activity dramatically alters the mRNA levels of over 1200 genes involved in diverse biological functions. These results, combined with the previously reported list of differentially expressed genes in mouse ES cells knocked down for Dot1L, demonstrates surprising cell type and species conservation of Dot1L-dependent gene expression. In 53Bp1⁻/⁻ cells, over 300 genes, many with functions in immune responses and apoptosis, were differentially expressed. To date, this is the first global analysis of gene expression in a 53Bp1-deficient cell line.
  
  
  CONCLUSIONS
  Taken together, our results uncover a negative role for Dot1L and H3K79 methylation in the DNA damage response in the absence of 53Bp1. They also enlighten the roles of Dot1L and 53Bp1 in gene expression and the control of DNA double-strand repair pathways in the context of chromatin.
authors.affiliation: Genome Stability Laboratory, School of Natural Sciences, Centre for Chromosome Biology, National University of Ireland Galway, Galway, Ireland.
authors.forename: Jennifer
authors.initials: J
authors.lastname: FitzGerald
authors.forename: Sylvie
authors.initials: S
authors.lastname: Moureau
authors.forename: Paul
authors.initials: P
authors.lastname: Drogaris
authors.forename: Enda
authors.initials: E
authors.lastname: O'Connell
authors.forename: Nebiyu
authors.initials: N
authors.lastname: Abshiru
authors.forename: Alain
authors.initials: A
authors.lastname: Verreault
authors.forename: Pierre
authors.initials: P
authors.lastname: Thibault
authors.forename: Muriel
authors.initials: M
authors.lastname: Grenon
authors.forename: Noel F
authors.initials: NF
authors.lastname: Lowndes
mesh_terms: D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000378:metabolism; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D000076983:Histone Methyltransferases; D011495:Histone-Lysine N-Methyltransferase / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D006657:Histones / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D046228:Microarray Analysis; D015398:Signal Transduction / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D000071857:Tumor Suppressor p53-Binding Protein 1
chemical_list: D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D006657:Histones; C498302:Trp53bp1 protein, mouse; D025521:Tumor Suppressor Proteins; D000071857:Tumor Suppressor p53-Binding Protein 1; C481527:Dot1l protein, mouse; D000076983:Histone Methyltransferases; D008780:Methyltransferases; D011495:Histone-Lysine N-Methyltransferase; D008239:Lysine